Functional changes in human leukemic cell line HL-60. A model for myeloid differentiation

Polar solvents induce terminal differentiation in the human promyelocytic leukemia cell line HL-60. The present studies describe the functional changes that accompany the morphologic progression from promyelocytes to bands and poly-morphonuclear leukocytes (PMN) over 9 d of culture in 1.3 percent dimethylsulfoxide (DMSO). As the HL-60 cells mature, the rate of O(2-) production increase 18-fold, with a progressive shortening of the lag time required for activation. Hexosemonophosphate shunt activity rises concomitantly. Ingestin of paraffin oil droplets opsonized with complement or Ig increases 10-fold over 9 d in DMSO. Latex ingestion per cell by each morphologic type does not change significantly, but total latex ingestion by groups of cells increases with the rise in the proportion of mature cells with greater ingestion capacities. Degranulation, as measured by release of β-glucuronidase, lysozyme, and peroxidase, reaches maximum after 3-6 d in DMSO, then declines. HL-60 cells contain no detectable lactoferrin, suggesting that their secondary granules are absent or defective. However, they kill staphylococci by day 6 in DMSO. Morphologically immature cells (days 1-3 in DMSO) are capable of O(2-) generation, hexosemonophosphate shunt activity, ingestion, degranulation, and bacterial killing. Maximal performance of each function by cells incubated in DMSO for longer periods of time is 50-100 percent that of normal PMN. DMSO- induced differentiation of HL-60 cells is a promising model for myeloid development.     

Enhancement and phototransduction in the ventral eye of limulus

Limulus ventral photoreceptors were voltage clamped to the resting (dark) potential and stimulated by a 20-ms test flash and a 1-s conditioning flash. At a constant level of adaptation, we measured the response to the test flash given in the dark (control) and the incremental response produced when the test flash occurred within the duration of the conditioning flash. The incremental response is defined as the response to the conditioning and test flashes minus the response to the conditioning flash given alone. When the test flash was presented within 100 ms after the onset of the conditioning flash we observed that: (a) for dim conditioning flashes the incremental response equaled the control response; (b) for intermediate intensity conditioning flashes the incremental response was greater than the control response (we refer to this as enhancement); (c) for high intensity conditioning flashes the incremental response nearly equaled the control response. Using 10-μm diam spots of illumnination, we stimulated two spatially separate regions of one photoreceptor. When the test flash and the conditioning flash were presented to the same region, enhancement was present; but when the flashes were applied to separate regions, enhancement was nearly absent. This result indicates that enhancement is localized to the region of illumination. We discuss mechanisms that may account for enhancement.     

Spinal p38beta isoform mediates tissue injury-induced hyperalgesia and spinal sensitization

Antagonist studies show that spinal p38 mitogen-activated protein kinase plays a crucial role in spinal sensitization. However, there are two p38 isoforms found in spinal cord and the relative contribution of these two to hyperalgesia is not known. Here we demonstrate that the isoforms are distinctly expressed in spinal dorsal horn: p38alpha in neurons and p38beta in microglia. In lieu of isoform selective inhibitors, we examined the functional role of these two individual isoforms in nociception by using intrathecal isoform-specific antisense oligonucleotides to selectively block the expression of the respective isoform. In these rats, down-regulation of spinal p38beta, but not p38alpha, prevented nocifensive flinching evoked by intraplantar injection of formalin and hyperalgesia induced by activation of spinal neurokinin-1 receptors through intrathecal injection of substance P. Both intraplantar formalin and intrathecal substance P produced an increase in spinal p38 phosphorylation and this phosphorylation (activation) was prevented when spinal p38beta, but not p38alpha, was down-regulated. Thus, spinal p38beta, probably in microglia, plays a significant role in spinal nociceptive processing and represents a potential target for pain therapy.     

The cDNA cloning and molecular evolution of reptile and pigeon lactate dehydrogenase isozymes

The cDNAs encoding lactate dehydrogenase isozymes LDH-A (muscle) and LDH-B (heart) from alligator and turtle and LDH-A, LDH-B, and LDH-C (testis) from pigeon were cloned and sequenced. The evolutionary relationships among vertebrate LDH isozymes were analyzed. Contrary to the traditional belief that the turtle lineage branched off before the divergence between the lizard/alligator and bird lineages, the turtle lineage was found to be clustered with either the alligator lineage or the alligator-bird clade, while the lizard lineage was found to have branched off before the divergence between the alligator/turtle and bird lineages. The pigeon testicular LDH-C isozyme was evidently duplicated from LDH-B (heart), so it is not orthologous to the mammalian testicular LDH-C isozymes.      

Disease resistance conferred by expression of a gene encoding H2O2-generating glucose oxidase in transgenic potato plants.

Plant defense responses to pathogen infection involve the production of active oxygen species, including hydrogen peroxide (H2O2). We obtained transgenic potato plants expressing a fungal gene encoding glucose oxidase, which generates H2O2 when glucose is oxidized. H2O2 levels were elevated in both leaf and tuber tissues of these plants. Transgenic potato tubers exhibited strong resistance to a bacterial soft rot disease caused by Erwinia carotovora subsp carotovora, and disease resistance was sustained under both aerobic and anaerobic conditions of bacterial infection. This resistance to soft rot was apparently mediated by elevated levels of H2O2, because the resistance could be counteracted by exogenously added H2O2-degrading catalase. The transgenic plants with increased levels of H2O2 also exhibited enhanced resistance to potato late blight caused by Phytophthora infestans. The development of lesions resulting from infection by P. infestans was significantly delayed in leaves of these plants. Thus, the expression of an active oxygen species-generating enzyme in transgenic plants represents a novel approach for engineering broad-spectrum disease resistance in plants.     

Characterization of a Nonparasitic Isolate of Bursaphelenchus xylophilus

Bursaphelenchus xylophilus isolate MPSy-1av was subcultured from pathotype MPSy-1. MPSy-1av is nonparasitic and does not establish in Pinus sylvestris, P. strobus, P. nigra, or P. taeda. This isolate produces ethanol as an end product of carbohydrate metabolism, whereas its parent pathotype, MPSy-1, does not. Alcohol dehydrogenase activity was easily detectable in homogenates of MPSy-1av but barely detectable in some homogenates of MPSy-1. Genomic differences were seen between MPSy-1 and M PSy-1av by restriction endonuclease analysis of total nematode DNA, and hybridization of DNA fragments to the alcohol dehydrogenase gene from Drosophila.